Roles for Transcription in Antibody Diversity

National Institute On Aging

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Abstract

To adapt this protocol for B cells, cells were fixed prior to nuclear isolation, and several critical adjustments were introduced to the procedure and reagents. We measured binding of H3K4me3 histone and RNA Polymerase II, detecting robust peaks with as little as 100k nuclei. Additionally, freeze-thaw of B cells prior to processing did not affect results, emphasizing the flexibility of this modified technique. Our modified protocol will allow one to quantify low abundance, non-histone proteins bound to DNA from limited numbers of B cells with more efficiency than can be achieved from the current standard, ChIP-seq.